The invention relates to a method for the determination of the activity of plasminogen activators in plasma or other biological fluids.
The key enzymes in the physiological fibrinolysis in mammalian blood are the two plasminogen activators a) tissue plasminogen activator (t-PA) and b) urinary plasminogen activator (u-PA). Plasminogen activators convert the inactive zymogen plasminogen into the protease plasmin. Plasmin degrades insoluble fibrin into soluble fibrin split products.
The activity of these PAs in human blood is of great diagnostic relevance for measuring the fibrinolytic potential of a patient. The fibrinolytic potential provides information on, inter alia, possible thrombo-embolisms which are to be expected. However, PAs occur only in small amounts (ng/ml) in plasma. This makes great demands on the sensitivity and specificity of the detection method.
The functional determination of PAs according to the state of the art entails their natural substrate, plasminogen, and a chromogenic plasmin substrate being added to the sample. In this case, the measured plasmin activity represents PA activity which is enhanced in direct proportion.
However, the plasmin which is produced is immediately inhibited, almost quantitatively, by alpha-2-antiplasmin which is likewise contained in the sample, so that only low plasmin activity can be measured.
The accuracy of functional PA determination thus depends on eliminating this antiplasmin effect. The methods for the functional determination of PAs described in the state of the art therefore either provide for additional plasma separation steps such as euglobulin precipitation or require acidification and subsequent neutralization of a highly diluted sample. Both methods are relatively time-consuming and methodologically demanding.